Wehner G, Friedmann B, Zimmermann U
Lehrstuhl für Biotechnologie, Würzburg, F.R.G.
Biochim Biophys Acta. 1990 Aug 24;1027(2):105-15. doi: 10.1016/0005-2736(90)90073-w.
Charge pulse experiments performed on the peat-bog alga Eremosphaera viridis revealed an unusual voltage relaxation behaviour. Injection of charge pulses of 1 microseconds duration resulted in an immediate charging of the membranes (time constant of the order of 40 ns). Nevertheless, the potential-measuring microelectrode recorded an exponential increase in membrane voltage with a time constant of about 1.3 ms. The maximum voltage value was recorded after about 3 ms, followed by an exponential decay with a time constant of about 9.6 ms. This biphasic time course was independent of the amplitude of the injected charge and of the location of the impaled microelectrodes in the vacuole. Centrifuged cells in which the chloroplasts and the other organelles were pelleted in one part of the cells showed the same electrical response. Electrical breakdown of the cell membranes resulted in the disappearance of the biphasic voltage response. In this case only the decaying relaxation process could be recorded with a time constant of 3 ms. After resealing of the membranes the original biphasic relaxation response was restored. Increasing concentrations of KCl in the bathing medium reduced both time constants almost correspondingly. The experimental findings were evaluated with an electrical equivalent circuit. Theoretical analysis with reference to the experimental data suggested that the delayed voltage response of the potential-recording electrode resulted from a membrane seal across the tip of this electrode. The resistance of this seal was calculated to be about 400 M omega. The specific resistances and capacitances of tonoplast and plasmalemma membranes were calculated from the decaying part of the biphasic relaxation curves. The average values were found to be 2.58 omega.m2 and 5 mF.m-2. The investigations reported here suggest that charge pulse experiments can be generally used for the detection of membrane and cytoplasmic material clogging of the tip of intracellular microelectrodes, a problem with which most electrophysiologists are faced when interpreting data obtained from impaled microelectrodes.
对泥炭沼泽藻类绿裸藻进行的充电脉冲实验揭示了一种不寻常的电压弛豫行为。注入持续时间为1微秒的充电脉冲会导致膜立即充电（时间常数约为40纳秒）。然而，电位测量微电极记录到膜电压呈指数增加，时间常数约为1.3毫秒。约3毫秒后记录到最大电压值，随后以约9.6毫秒的时间常数呈指数衰减。这种双相时间进程与注入电荷的幅度以及刺入液泡中的微电极位置无关。叶绿体和其他细胞器在细胞的一部分中沉淀的离心细胞显示出相同的电反应。细胞膜的电击穿导致双相电压反应消失。在这种情况下，只能记录到时间常数为3毫秒的衰减弛豫过程。膜重新密封后，恢复了原来的双相弛豫反应。浴液中氯化钾浓度的增加几乎相应地降低了两个时间常数。用电等效电路对实验结果进行了评估。参照实验数据进行的理论分析表明，电位记录电极的延迟电压反应是由该电极尖端的膜密封引起的。计算得出该密封的电阻约为400兆欧。根据双相弛豫曲线的衰减部分计算了液泡膜和质膜的比电阻和电容。发现平均值分别为2.58欧·平方米和5毫法·平方米。此处报道的研究表明，充电脉冲实验通常可用于检测细胞内微电极尖端的膜和细胞质物质堵塞，这是大多数电生理学家在解释从刺入微电极获得的数据时面临的一个问题。
